Mycoplasma genitalium (MG) is an emerging human pathogen increasingly recognized for its etiologic role in reproductive tract disease in men and women and its ability to persist for months and even years in vivo despite the induction of specific antibodies during infection. Antigenic variation of the immunodominant surface proteins, MgpB and MgpC, is thought to be critical for the ability of MG to evade the host immune response and persist. This RecA-mediated process is accomplished by reciprocal segmental recombination between the mgpBC expression site and architecturally distinct archived homeologous (partially homologous) truncated sequences (termed MgPars) distributed throughout the chromosome. In addition to antigenic variation, MgpB and MgpC can undergo phase variation, the focus of the current proposal. Phase variants are unable to adhere to host cells and can be easily detected by the hemadsorption negative [HA(-)] phenotype of colonies cultured on agar plates. Only two phase variants have been characterized to date; both contain large deletions in the mgpBC expression site, the adjacent MgPar site, and the intervening sequences, and thus are irreversible. Phase variants, in which the mgpBC expression site sequences were reciprocally exchanged with the MgPar sequences, were initially detected by PCR, but were not characterized further. However, the unusual architecture of the resulting sequences predicted that they could revert to wild type by the reverse of the recombination event that generated them, suggesting their relevance in pathogenesis. Several phase variants have now been isolated and sequenced, confirming their reciprocal and reversible nature. These findings, and the detection of phase variants at a frequency higher than that of antigenic variants, led to the hypotheses that phase variants are critical to the biology of MG and arise by a novel method of reversible reciprocal recombination dependent on the unusual differential architecture of the mgpBC and MgPar sites. The current study will explore the hypotheses that phase variants (1) arise via a mechanism not found in other bacteria, (2) are selected by antibodies targeting the conserved regions of MgpB, and (3) are important for the evasion of the immune response in vitro and in vivo. These hypotheses will be explored by (1) evaluating the architecture and reversibility of spontaneous phase variants obtained in vitro, (2) determining if rabbit antibodies targeting a conserved (nonvariable) immunodominant region of MgpB select for phase variants in vitro, and (3) assessing the in vivo significance of immune evasion by enumerating phase variants in longitudinally collected archived genital specimens from a primate model of MG infection and the ability of archived primate sera to select for phase variants in vitro. This study will employ innovative techniques to reveal the mechanisms responsible for this novel system of phase variation in an extremely fastidious pathogen with a very limited genome (580 kb) and few identified recombination genes. In addition, the biologic consequences of phase variation will be revealed for the first time and may lead to future studies of novel methods of prevention and treatment.